Nitroglycerin (glyceryl trinitrate, GTN) has been used to treat angina pectoris and congestive heart failure for over 130 years. It is generally accepted that GTN is converted in vascular smooth muscle cells to nitric oxide (NO) or an NO congener (S-nitrosothiol, SNO), which activates guanylate cyclase and this relaxes vascular smooth muscle. However, the molecular mechanism of GTN biotransformation has remained a mystery, and it is not understood why therapy with GTN is associated with nitrate tolerance (loss of clinical sensitivity to GTN) and oxidative stress (caused by GTN being an oxidant). Nitrate tolerance from GTN administration is especially evident when GTN is administered intravenously in the treatment of unstable angina and heart failure in the form of a conventional composition containing 20 mM GTN in 50% ethanol (e.g. 7 to 8 molar ethanol or over 100 times the moles of GTN) in a dosage of GTN ranging from 5 to 100 μg/min, e.g. from 20-54 μg/min, for 40 to 100 minutes. Nitrate tolerance is also noted when GTN is administered orally or topically and when isorbide dinitrate is administered orally or topically, in the treatment of angina and heart failure. Currently, nitrate tolerance is treated by increasing the dosage of nitrate administered, and this works for a while but not over the long term or for a chronic disorder (e.g., endothelulial dysfunction ensues).
GTN biotransformation has previously been found to be tissue and cell specific and to yield 1,2-glyceryl dinitrate (1,2-GDN), 1,3-glyceryl dinitrate (1,3-GDN), inorganic nitrite and NO (or SNO) in differing amounts and, ratios. Published data indicates that in vascular smooth muscle cells, 1,2-GDN is the predominant dinitrate metabolite, and either NO or SNO mediates vasodilation. Furthermore, published data indicates that the vasorelaxation-dependent formation of 1,2-GDN is reduced in GTN-tolerant blood vessels. These data suggest that the enzyme(s) generating NO bioactivity from GTN catalyzes the selective formation of 1,2-GDN (over 1,3-GDN) and that loss of this activity at least partly accounts for tolerance. Several mechanisms of vascular smooth muscle relaxation by GTN have been proposed; enzymes proposed for mediating GTN metabolism include glutathione-S-transferases, cytochrome P450 reductase, cytochrome P450, xanthine oxide reductase and an unidentified microsomal protein. However, none of these enzymes catalyzes the selective formation of 1,2-GDN or is inhibited in tolerant vessels, so none of these enzymes can be the enzyme or enzymes mediating GTN metabolism.